Human RIG-I Antibody

Detection of Human RIG-I by Western Blot.

Western blot shows lysates of A172 human glioblastoma cell line and Nalm-6 human Pre-B acute lymphocytic leukemia cell line. PVDF membrane was probed with 1 µg/mL of Goat Anti-Human RIG-I Antigen Affinity-purified Polyclonal Antibody (Catalog # AF4859) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF019). A specific band was detected for RIG-I at approximately 105 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 8.

Detection of RIG-I by Western Blot

RuV infection induces STAT1 activation and ISG expression. (A) Protein lysates from U373MG cells (a) or CCF-STTG1 cells (b) that were uninfected (M) or infected with RuV RA27/3 strain (+) at 2 and 4 days postinfection (dpi) (MOI of 1.8) were analyzed by immunoblotting for STAT1, Tyr701-phosphorylated STAT1 (p-STAT1), and RuV-encoded capsid protein. GAPDH protein expression was analyzed as a reference. Densitometric analysis of the capsid protein was performed with NIH ImageJ software. Signal intensity of the capsid protein was normalized to that of GAPDH, then shown as a relative ratio of U373MG cells infected with RuV at 2 dpi. (B) STAT1-EGFP fusion protein (green) translocated to the nucleus was detected in RuV-infected U373MG-STAT1-EGFP cells at 2 dpi by fluorescence microscopy. Typical cells positive for nuclear localization of EGFP fusion protein are shown. Nuclei were counterstained with Hoechst 33342 (blue). Images were acquired at a magnification of 40× objective lens. (C) Total RNAs extracted from U373MG cells infected with RuV RA27/3 strain (+) or heat-inactivated RuV RA27/3 strain (HI) or uninfected (M) at 2 and 4 dpi were subjected to RT-PCR for the detection of expression of MDA5, ISG56, OAS1, MX1, and viperin mRNAs. TBP mRNA was analyzed as a reference. The PCR products were analyzed by agarose gel electrophoresis. (D) Protein lysates from U373MG cells (a) or CCF-STTG1 cells (b) that were uninfected (M) or infected with RuV RA27/3 strain (+) at 2 and 4 dpi (MOI of 1.8) were analyzed by immunoblotting for the protein expression of ISGs (RIG-I, MDA5, ISG56, MX1, and viperin). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) protein was analyzed as a reference. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/36077193), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of RIG-I by Western Blot

Short hairpin RNA-mediated MAVS knockdown attenuated the RuV-triggered IFN-beta signaling pathway. (A) Total RNAs extracted from U373MG cells expressing MAVS-targeting shRNAs (sh1MAVS and sh2MAVS) or a non-targeting shRNA (shNT) that were uninfected (M) or infected with the RuV RA27/3 strain (+) at 2 dpi (MOI of 1.8) were subjected to RT-PCR for the detection of MAVS, IFN-beta, and viperin mRNAs and RuV RNAs encoding the capsid protein. TBP mRNA was analyzed as a reference. The products were analyzed by agarose gel electrophoresis. (B) Protein lysates from shRNAs (sh1MAVS, sh2MAVS, and shNT)-expressing U373MG cells that were uninfected (M) or infected with the RuV RA27/3 strain (+) at 2 dpi (MOI of 1.8) were analyzed by immunoblotting for the protein expression of MAVS, STAT1, Tyr701-phosphorylated STAT1 (p-STAT1), ISGs (RIG-I, ISG56, MX1, and viperin), and RuV-encoded capsid proteins. GAPDH protein was analyzed as a reference. Densitometric analysis of MAVS and capsid proteins was performed with NIH ImageJ software. Signal intensity of those proteins was normalized to that of GAPDH, then shown as a relative ratio of shNT-expressing U373MG cells infected with the RuV RA27/3 strain. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/36077193), licensed under a CC-BY license. Not internally tested by R&D Systems.